1. Field of the Invention
The present invention relates to the field of molecular biology, and in particular, to the expression of heterologous biomass degrading enzymes in gram-positive thermophilic anaerobic bacteria.
2. Background Art
Thermophilic microorganisms are useful for a variety of industrial processes. For example, thermophilic microorganisms can be used as biocatalysts in reactions at higher operating temperatures than can be achieved with mesophilic microorganisms. Thermophilic organisms are particularly useful in biologically mediated processes for energy conversion, such as the production of ethanol from plant biomass, because higher operating temperatures allow more convenient and efficient removal of ethanol in vaporized form from the fermentation medium.
The ability to metabolically engineer thermophilic microorganisms to improve various properties (e.g., ethanol production, breakdown of lignocellulosic materials), would allow the benefit of higher operating temperatures to be combined with the benefits of using industrially important enzymes from a variety of sources in order to improve efficiency and lower the cost of production of various industrial processes, such as energy conversion and alternative fuel production.
Thermophilic anaerobic gram-positive bacteria such as Thermoanaerobacterium saccharolyticum (“T. sacch”) can be particularly useful in methods of energy conversion since they can grow at temperatures above 40° C. and are readily able to utilize cellobiose (a disaccharide) and xylose (a monosaccharide) as energy sources. However, T. sacch are not able to hydrolyze cellulose efficiently. Previous experiments have demonstrated that it is possible to expresses heterologous cellulases in T. sacch (Mai and Wiegel, Applied and Environmental Microbiology, 66: 4817-4821 (2000)). However, major shortcomings of the previous strains were that ethanol was not the sole metabolic product and that they were unable to achieve sufficient levels of secreted enzyme to be industrially useful. In contrast, the present invention provides T. sacch and other transformed thermophilic anaerobic bacteria that express high levels of heterologous cellulases that can efficiently produce ethanol or other useful fermentation products such as lactic acid, acetic acid, or CO2 from cellulose.